Calcium montmorillonite clay reduces urinary biomarkers of fumonisin B₁ exposure in rats and humans

Abstract

Fumonisin B₁ (FB₁) is often a co-contaminant with aflatoxin (AF) in grains and may enhance AF's carcinogenicity by acting as a cancer promoter. Calcium montmorillonite (i.e. NovaSil, NS) is a possible dietary intervention to help decrease chronic aflatoxin exposure where populations are at risk. Previous studies show that an oral dose of NS clay was able to reduce AF exposure in a Ghanaian population. In vitro analyses from our laboratory indicated that FB₁ (like aflatoxin) could also be sorbed onto the surfaces of NS. Hence, our objectives were to evaluate the efficacy of NS clay to reduce urinary FB₁ in a rodent model and then in a human population highly exposed to AF. In the rodent model, male Fisher rats were randomly assigned to either FB₁ control, FB₁ + 2% NS or absolute control group. FB₁ alone or with clay was given as a single dose by gavage. For the human trial, participants received NS (1.5 or 3 g day⁻¹) or placebo (1.5 g day⁻¹) for 3 months. Urines from weeks 8 and 10 were collected from the study participants for analysis. In rats, NS significantly reduced urinary FB₁ biomarker by 20% in 24 h and 50% after 48 h compared to controls. In the humans, 56% of the urine samples analysed (n = 186) had detectable levels of FB₁. Median urinary FB₁ levels were significantly (p < 0.05) decreased by >90% in the high dose NS group (3 g day⁻¹) compared to the placebo. This work indicates that our study participants in Ghana were exposed to FB₁ (in addition to AFs) from the diet. Moreover, earlier studies have shown conclusively that NS reduces the bioavailability of AF and the findings from this study suggest that NS clay also reduces the bioavailability FB₁. This is important since AF is a proven dietary risk factor for hepatocellular carcinoma (HCC) in humans and FB₁ is suspected to be a dietary risk factor for HCC and oesophageal cancer in humans.

Figures

Figure 1

Comparison of FB1 with OPA derivatisation vs. NDA derivatisation. Both OPA and NDA derivatisation products were linear within a range of 483–4.83 ng (A and B). The relative intensity of NDA derivatisation was 2–5 times higher than that of OPA (concentration dependent) allowing for a lower limit of detection (0.242 ng) while retaining linearity (C and D).

Figure 2

NovaSil reduction of total excreted and creatinine standardised urinary FB1 in Fischer 344 rats. Data represents the cycle of urinary FB1 excretion following 25mg kg_1 dose by gavage. Urinary FB1 was reduced by 27% in the NS treated group. This reduction was statistically significant (p_0.05) when NS treated and non-treated groups were compared using the Kruskal–Wallis and Wilcoxon rank-sum test. Data represents the means and standard error. Non-detectable levels of FB1 were documented for the control group at all time points.

Figure 3

Distribution of urinary FB1 between treatment groups at weeks 8 and 10. Data represents the distribution of FB1 (ng FB1 ml_1 urine) in the various treatment groups based on a NovaSil intervention in Ghana. There was a statistically significant difference among treatments (p ≤ 0.05). There was a significant difference between the median FB1 levels in the NS treated groups vs. the placebo group (p ≤ 0.05). There was not a significant difference between median FB1 levels between the high and low dose groups (p = 0.566).

Figure 4

Distribution of creatinine standardised urinary FB1 between treatment groups at weeks 8 and 10. Data represents the distribution of FB1 (ng FB1 mg_1 creatinine) in the various treatment groups based on a NovaSil intervention in Ghana. There was a statistically significant difference among treatments (p ≤ 0.05). There was a significant difference between the median FB1 levels in the NS treated groups vs. the placebo group (p ≤ 0.05). There was not a significant difference between median FB1 levels in the high and low dose groups (p = 0.955).

Figure 5

MALDI-TOF mass spectra (MS). MS are based on relative intensity (y-axis) of depicted peaks (highest peak = 100%). (A) MS of underivatised standard FB1 in water (723 m/z). (B) MS of NDA derivatised FB1 from spiked urine sample (~906.1 m/z) and NDA derivative + Na ion (~928 m/z). (C) MS of a representative FB1 positive participant urine sample verifying presence of FB1.